LOCUS       sequence-78194-e        7681 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Pyronic, FRET sensor that allow quantitative measurement of pyruvate
            transport, pyruvate production and mitochondrial pyruvate 
            consumption in intact individual cells at high temporal resolution..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7681)
  AUTHORS   San Martin A, Ceballo S, Baeza-Lehnert F, Lerchundi R, Valdebenito 
            R, Contreras-Baeza Y, Alegria K, Barros LF
  TITLE     Imaging Mitochondrial Flux in Single Cells with a FRET Sensor for 
            Pyruvate.
  JOURNAL   PLoS One. 2014 Jan 21;9(1):e85780. doi: 
            10.1371/journal.pone.0085780. eCollection 2014 Jan 21.
  PUBMED    24465702
REFERENCE   2  (bases 1 to 7681)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "PLoS One.";
            date: "2014-01-21"; volume: "9(1):e85780. doi"; pages: " 
            10.1371/journal.pone.0085780. eCollection 2014 Jan 21"
COMMENT     Sequence Label: Pyronic/pcDNA3.1(-)
FEATURES             Location/Qualifiers
     source          1..7681
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             5..712
                     /codon_start=1
                     /product="monomeric teal (cyan) fluorescent protein (Ai et 
                     al., 2006)"
                     /label=mTFP1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEETTMGVIKPDMKIKLKMEGNVNGHAFVIEGEGEGKPYDG
                     TNTINLEVKEGAPLPFSYDILTTAFAYGNRAFTKYPDDIPNYFKQSFPEGYSWERTMTF
                     EDKGIVKVKSDISMEEDSFIYEIHLKGENFPPNGPVMQKKTTGWDASTERMYVRDGVLK
                     GDVKHKLLLEGGGHHRVDFKTIYRAKKAVKLPDYHFVDHRIEILNHDKDYNKVTVYESA
                     VARNSTDGMDELYK"
     protein_bind    725..749
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(1547..1549,1550..1552,1553..2266)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1592..1613)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1853..1872)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2200..2221
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2293..2310)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2299..2523
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2569..2997
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2656..2675)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2866..2887
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3006..3026)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3011..3340
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3191..3326
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3253..3272
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3407..4201
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3461..3480)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4071..4090
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4375..4496
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4412..4431)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4466..4485
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4545..4561)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4545..4561)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4558..4580)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4569..4585
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4593..4599,4600..4617,4618..4623))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4638..4659
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4776..4793)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4947..5532)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5027..5046)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5703..6494,6495..6563))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6326..6345
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6564..6668)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6743..6762)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        6934..7313
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7314..7517
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7468..7488
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     7562..7581
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7562..7580
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gaacatggtg agcaagggcg aggagaccac aatgggcgta atcaagcccg acatgaagat
       61 caagctgaag atggagggca acgtgaatgg ccacgccttc gtgatcgagg gcgagggcga
      121 gggcaagccc tacgacggca ccaacaccat caacctggag gtgaaggagg gagcccccct
      181 gcccttctcc tacgacattc tgaccaccgc gttcgcctac ggcaacaggg ccttcaccaa
      241 gtaccccgac gacatcccca actacttcaa gcagtccttc cccgagggct actcttggga
      301 gcgcaccatg accttcgagg acaagggcat cgtgaaggtg aagtccgaca tctccatgga
      361 ggaggactcc ttcatctacg agatacacct caagggcgag aacttccccc ccaacggccc
      421 cgtgatgcag aagaagacca ccggctggga cgcctccacc gagaggatgt acgtgcgcga
      481 cggcgtgctg aagggcgacg tcaagcacaa gctgctgctg gagggcggcg gccaccaccg
      541 cgttgacttc aagaccatct acagggccaa gaaggcggtg aagctgcccg actatcactt
      601 tgtggaccac cgcatcgaga tcctgaacca cgacaaggac tacaacaagg tgaccgttta
      661 cgagagcgcc gtggcccgca actccaccga cggcatggac gagctgtaca agagatctgg
      721 taccacaagt ttgtacaaaa aagcaggctc cgaattcgcc cttggtacca tggcctacag
      781 caaaatccgc caaccaaaac tctccgatgt gattgagcag caactggagt ttttgatcct
      841 cgaaggcact ctccgcccgg gcgaaaaact cccaccggaa cgcgaactgg caaaacagtt
      901 tgacgtctcc cgtccctcct tgcgtgaggc gattcaacgt ctcgaagcga agggcttgtt
      961 gcttcgtcgc cagggtggcg gcacttttgt ccagagcagc ctatggcaaa gtttcagcga
     1021 tccgctggtg gagctgctct ccgaccatcc tgagtcacag tatgacttgc tcgaaacacg
     1081 acacgccctg gaaggtatcg ccgcttatta cgccgcgctg cgtagtaccg atgaagacaa
     1141 ggaacgcatc cgtgaactcc accacgccat agagctggcg cagcagtctg gcgatctgga
     1201 cgcggaatca aacgccgtac tccagtatca gattgccgtc accgaagcgg cccacaatgt
     1261 ggttctgctt catctgctaa ggtgtatgga gccgatgttg gcccagaatg tccgccagaa
     1321 cttcgaattg ctctattcgc gtcgcgagat gctgccgctg gtgagtagtc accgcacccg
     1381 catatttgaa gcgattatgg ccggtaagcc ggaagaagcg cgcgaagcat cgcatcgcca
     1441 tctggccttt atcgaagaaa ttttgctcga cagaagtcgt gaagagagcc gccgtgagcg
     1501 ttctctgcgt cgtctggagc aacgaaagaa tcttaagaga tctaccatgg tgagcaaggg
     1561 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
     1621 ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct
     1681 gaagctgatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct
     1741 gggctacggc ctgcagtgct tcgcccgcta ccccgaccac atgaagcagc acgacttctt
     1801 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg
     1861 caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga
     1921 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa
     1981 ctacaacagc cacaacgtct atatcaccgc cgacaagcag aagaacggca tcaaggccaa
     2041 cttcaagatc cgccacaaca tcgaggacgg cggcgtgcag ctcgccgacc actaccagca
     2101 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagctacca
     2161 gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
     2221 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtaaaagc ttaagtttaa
     2281 accgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc
     2341 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg
     2401 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg
     2461 acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta
     2521 tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta
     2581 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     2641 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     2701 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     2761 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     2821 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     2881 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     2941 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat
     3001 tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
     3061 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
     3121 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
     3181 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     3241 actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa
     3301 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat
     3361 atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga
     3421 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc
     3481 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc
     3541 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc
     3601 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac
     3661 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc
     3721 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac
     3781 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg
     3841 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct
     3901 cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt
     3961 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg
     4021 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac
     4081 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg
     4141 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg
     4201 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     4261 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     4321 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     4381 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     4441 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     4501 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     4561 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     4621 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     4681 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     4741 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     4801 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     4861 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     4921 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     4981 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     5041 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     5101 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     5161 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     5221 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     5281 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     5341 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     5401 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     5461 aaacaaacca ccgctggtag cggttttttt gtttgcaagc agcagattac gcgcagaaaa
     5521 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     5581 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     5641 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     5701 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     5761 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     5821 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     5881 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     5941 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     6001 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     6061 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     6121 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     6181 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     6241 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     6301 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     6361 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     6421 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     6481 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     6541 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     6601 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     6661 gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg
     6721 atcccctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc
     6781 tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa
     6841 caaggcaagg cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc
     6901 tgcttcgcga tgtacgggcc agatatacgc gttgacattg attattgact agttattaat
     6961 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
     7021 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
     7081 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
     7141 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
     7201 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat
     7261 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
     7321 ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
     7381 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
     7441 aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg
     7501 tctatataag cagagctctc tggctaacta gagaacccac tgcttactgg cttatcgaaa
     7561 ttaatacgac tcactatagg gagacccaag ctggctagcg tttaaacggg ccctctagac
     7621 tcgagcggcc gccactgtgc tggatatctg cagaattcca ccacactgga ctagtggatc
     7681 c
//
